Alzheimer's disease is characterized pathologically by abnormal depositions of ? amyloid proteins in the brain, called senile plaques. Unfortunately, the pathophysiological role of the ?-amyloid peptides in causing Alzheimer's disease and how senile plaques affect the neuronal membrane is still unknown. Although many findings strongly indicate that the pathophysiological effects of amyloid plaques may partially be caused by direct interactions between membrane lipids and ?-amyloid peptides, the molecular basis of the interaction is still not well understood. According to one hypothesis, the ?-amyloid peptides interact electrostatically with the neuronal membrane, while according to another, the interaction is hydrophobic such that the peptide penetrates the membrane and disrupts its functionality. Furthermore, a great number of studies indicate that cholesterol or cholesterol metabolism may be important in the pathophysiology of Alzheimer's disease. Therefore, it would be very important to elucidate how depletion or enrichment of cholesterol relates to Alzheimer's disease. The main goal in this project is to elucidate the interaction of the ?-amyloid peptide (25- 35) and phospholipid model membranes with and without cholesterol by using new electron paramagnetic resonance (EPR) methods developed in our laboratory. The working hypothesis is that the hydrophobic part of ?AP(25-25) penetrates into the phospholipid bilayer and thus changes the fluidity of the bilayer, making the membrane more rigid, while at the same time due to the insertion of the peptide the permeability of the membrane increases to allow a deeper penetration of water and cations into the bilayer. These two interactions disrupt the phospholipid bilayer and can account for one of the possible mechanisms for ?AP(25-25) neurotoxicity. Cholesterol added to the phospholipid bilayer should reduce the insertion of ?AP(25-25) into the bilayer, and thus preserve bilayer integrity from the disrupting effect of the ?-amyloid peptide, thereby reducing the neurotoxicity of the peptide. To test the hypothesis, we propose conducting a wide range of electron paramagnetic resonance experiments on phospholipid vesicles, peptide-phospholipid vesicles, cholesterol-phospholipid vesicles and peptide-cholesterol-phospholipid vesicles. These experiments are designed to give a complete knowledge of the changes in membrane fluidity and permeability across the phospholipid bilayer, so that the location of the peptide in the phospholipid bilayer and the structural changes of the aggregate as a whole can be determined. This information may provide new insight into how the ?-amyloid peptide affects the neurodegenerative process associated with Alzheimer's disease.